DNA Damage Checkpoint Maintains Cdh1 in an Active State to Inhibit Anaphase Progression

Abstract

DNA damage checkpoint prevents segregation of damaged chromosomes by imposing cell-cycle arrest. In budding yeast, Mec1, Chk1, and Rad53 (homologous to human ATM/ATR, Chk1, and Chk2 kinases, respectively) are among the main effectors of this pathway. The DNA damage checkpoint is thought to inhibit chromosome segregation by preventing separase-mediated cleavage of cohesins. Here, we describe a regulatory network that prevents segregation of damaged chromosomes by restricting spindle elongation and acts in parallel with inhibition of cohesin cleavage. This control circuit involves Rad53, polo kinase, the anaphase-promoting complex activator Cdh1, and the bimC kinesin family proteins Cin8 and Kip1. The inhibition of polo kinase by Rad53-dependent phosphorylation prevents it from inactivating Cdh1. As a result, Cdh1 remains in a partially active state and limits Cin8 and Kip1 accumulation, thereby restraining spindle elongation. Hence, the DNA damage checkpoint suppresses both cohesin cleavage and spindle elongation to preserve chromosome stability.

“Reductional anaphase” in replication-defective cells is caused by ubiquitin-conjugating enzyme Cdc34-mediated deregulation of the spindle

“…It has been shown previously that both replication checkpoint and DNA damage checkpoint are capable of restraining spindle elongation. 13,25 However, our study strongly suggests that the task dilution), anti-G6PD polyclonal antibodies (Sigma, 1:10000 dilution), anti-Mad1 antibodies (a gift from Dr Kevin Hardwick) and anti-Rad53 (Santa Curz, 1:1000 dilution), respectively. Enhanced chemiluminescence kit from Santa Cruz was used for all western blot analyses according to the manufacturer's instructions.…”

“…13 The DNA damage checkpoint has also been reported to regulate mitotic spindle elongation (in parallel with the inhibition of cohesin cleavage) to prevent segregation of damaged chromosomes. 25 Hence, the regulation of spindle dynamics is an important element in the checkpoint control networks in budding yeast. In this study, we have investigated the mechanism underlying the segregation of unreplicated chromosomes in Cdc6-deficient cells.…”

When genome integrity and cell cycle decisions collide: roles of polo kinases in cellular adaptation to DNA damage

“…1a). In contrast, another study recently suggested that there was a Rad53-dependent reduction of Cdc5 activity by *50-66 % in the presence of DNA damage (Zhang et al 2009) and that this lowered Cdc5 activity might prevent normal mitotic progression by maintaining the APC activator Cdh1 (Zhang et al 2009) and the mitotic exit inhibitor Bfa1 in hypophosphorylated states (ValerioSantiago et al 2013) (Fig. 1b).…”

“…Many different proteins involved in various cellular processes have been connected to the adaptation response, including cell cycle regulators (Jin and Wang 2006;Toczyski et al 1997;Zhang et al 2009), chromatin-remodeling enzymes (Papamichos-Chronakis et al 2006), autophagy factors (Dotiwala et al 2013), and centrosome/spindle pole body effectors (Hu et al 2001;Valerio-Santiago et al 2013). How this group of apparently disparate proteins regulates the adaptation to DNA damage and whether they work together or in separate pathways to overcome checkpointinduced cell cycle arrest is still unclear.…”

“…1b). Since Cdh1 and Bfa1 are both inhibited by Cdc5 phosphorylation, it was proposed that lowering Cdc5 kinase activity would prevent cell cycle progression by limiting mitotic spindle elongation (a Cdh1 function; Zhang et al 2009) and preventing exit from mitosis (a Bfa1 function; Valerio-Santiago et al 2013).…”